Helper T lymphocytes (HTL) play several important functions in immunity to pathogens. Firstly, they provide help for induction of both CTL and antibody responses. By both direct contact and by secreting lymphokines such as IL2 and IL4, HTL promote and support the expansion and differentiation of T and B cell precursors into effector cells. In addition, HTL can also be effectors in their own right, an activity also mediated by direct cell contact and secretion of lymphokines, such as IFNxcex3 and TNFxcex1. HTL have been shown to have direct effector activity in case of tumors, as well as viral, bacterial, parasitic, and fungal infections.
HTL recognize a complex formed between Class II MHC molecules and antigenic peptides, usually between 10 and 20 residues long, and with an average size of between 13 and 16 amino acids. Peptide-Class II interactions have been analyzed in detail, both at the structural and functional level, and peptide motifs specific for various human and mouse Class II molecules have been proposed.
In the last few years, epitope based vaccines have received considerable attention as a possible mean to develop novel prophylactic vaccines and immunotherapeutic strategies. Selection of appropriate T and B cell epitopes should allow to focus the immune system toward conserved epitopes of pathogens which are characterized by high sequence variability (such as HIV, HCV and Malaria).
In addition, focusing the immune response towards selected determinants could be of value in the case of various chronic viral diseases and cancer, where T cells directed against the immunodominant epitopes might have been inactivated while T cells specific for subdominant epitopes might have escaped T cell tolerance. The use of epitope based vaccines also allows to avoid xe2x80x9csuppressivexe2x80x9d T cell determinants which induce TH2 responses, in conditions where a TH1 response is desirable, or vice versa.
Finally, epitope based vaccines also offer the opportunity to include in the vaccine construct epitopes that have been engineered to modulate their potency, either by increasing MHC binding affinity, or by alteration of its TCR contact residues, or both. Inclusion of completely synthetic non-natural or generically unrelated to the pathogen epitopes (such as TT derived xe2x80x9cuniversalxe2x80x9d epitopes), also represents a possible mean of modulating the HTL response toward a TH1, or TH2 phenotype.
Once appropriate epitope determinants have been defined, they can be assorted and delivered by various means, which include lipopeptides, viral delivery vectors, particles of viral or synthetic origin, naked or particle absorbed cDNA.
However, before appropriate epitopes can be defined, one major obstacle has to be overcome, namely the very high degree of polymorphism of the MHC molecules expressed in the human population. In fact, more than two hundred different types of HLA Class I and Class II molecules have already been identified. It has been demonstrated that in the case of HLA Class I molecules, peptides capable of binding several different HLA Class I molecules can be identified. Over 60% of the known HLA Class I molecules can, in fact, be grouped in four broad HLA supertypes, characterized by similar peptide binding specificities (HLA supermotifs).
In the case of Class III molecules, it is also known that peptides capable of binding multiple HLA types and of being immunogenic in the context of different HLA molecules do indeed exist. Until now, however, a general method for their identification has not been developed, probably at least in part a reflection of the fact that quantitative DR binding assays are labor intensive and that a large number of alleles [is?/needs?] to be considered.
The present invention addresses these and other needs.
The present invention is based, at least in part, on the discovery and validation of specific motifs and assay systems for various DR molecules, representative of the worldwide population. Their application to the identification of broadly degenerate HLA Class II binding peptides is also described.
Definitions
The term xe2x80x9cpeptidexe2x80x9d is used interchangeably with xe2x80x9coligopeptidexe2x80x9d in the present specification to designate a series of residues, typically L-amino acids, connected one to the other typically by peptide bonds between the alpha-amino and carbonyl groups of adjacent amino acids. The oligopeptides of the invention are less than about 50 residues in length and usually consist of between about 10 and about 30 residues, more usually between about 12 and 25, and often 15 and about 20 residues.
An xe2x80x9cimmunogenic peptidexe2x80x9d is a peptide which comprises an allele-specific motif such that the peptide will bind an MHC molecule and induce a HTL response. Immunogenic peptides of the invention are capable of binding to an appropriate HLA molecule and inducing HTL response against the antigen from which the immunogenic peptide is derived.
A xe2x80x9cconserved residuexe2x80x9d is a conserved amino acid occupying a particular position in a peptide motif typically one where the MHC structure may provide a contact point with the immunogenic peptide. One to three, typically two, conserved residues within a peptide of defined length defines a motif for an immunogenic peptide. These residues are typically in close contact with the peptide binding groove, with their side chains buried in specific pockets of the groove itself.
The term xe2x80x9cmotifxe2x80x9d refers to the pattern of residues of defined length, usually between about 8 to about 11 amino acids, which is recognized by a particular MHC allele.
The term xe2x80x9csupermotifxe2x80x9d refers to motifs that, when present in an immunogenic peptide, allow the peptide to bind more than one HLA antigen. The supermotif preferably is recognized by at least one HLA allele having a wide distribution in the human population, preferably recognized by at least two alleles, more preferably recognized by at least three alleles, and most preferably recognized by more than three alleles.
The phrases xe2x80x9cisolatedxe2x80x9d or xe2x80x9cbiologically purexe2x80x9d refer to material which is substantially or essentially free from components which normally accompany it as found in its native state. Thus, the peptides of this invention do not contain materials normally associated with their in situ environment, e.g., MHC I molecules on antigen presenting cells. Even where a protein has been isolated to a homogenous or dominant band, there are trace contaminants in the range of 5-10% of native protein which co-purify with the desired protein. Isolated peptides of this invention do not contain such endogenous co-purified protein.
The term xe2x80x9cresiduexe2x80x9d refers to an amino acid or amino acid mimetic incorporated in an oligopeptide by an amide bond or amide bond mimetic.